Immune suppressive product

ABSTRACT

An immune suppressive product prepared by injecting an allergen or a mixture of allergens into the body of milk-producing species. Said product being the milk or a polypeptide subfraction of milk obtained from the allergen treated host. The immune suppressive product(s) is milk and or the polypeptide fractions contained therein, which is ostensively free of the intact allergen or allergens used for the treatment of the host. The immune suppressive factor(s) being a subfraction of the allergen used for the treatment. A method of preparing immune suppressive polypeptides from intact allergens, which involves injection of the specific intact allergens into a milk-producing species, collecting the immune suppressive polypeptide fractions of the intact allergens from the milk of the treated host. The immune suppressive milk containing said polypeptide fractions, and/or the polypeptide fractions obtained from said milk, are nonreactive in animals and humans as allergens. Said factor(s), however, are highly effective in preventing or alleviating allergic reactions.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a division of application Ser. No. 07/815,630, filedDec. 30, 1991, which is a continuation-in-part of U.S. patentapplication Ser. No. 07/431,639, filed Nov. 6, 1989, now U.S. Pat. No.5,130,128 and a continuation-in-part of U.S. patent application Ser. No.07/177,223, filed Apr. 4, 1988 now U.S. Pat. No. 4,956,349; U.S.application Ser. No. 07/431,639 is a continuation-in-part application ofU.S. application Ser. No. 07/161,039, filed Feb. 26, 1988 now U.S. Pat.No. 4,879,110; both U.S. application Ser. No. 07/161,039 and U.S.application Ser. No. 07/177,223 are continuation-in-part applications ofU.S. patent application Ser. No. 07/001,848, filed Jan. 9, 1987, nowU.S. Pat. No. 4,897,265, which is a divisional of U.S. patentapplication Ser. No. 06/546,162, filed Oct. 27, 1983, now U.S. Pat. No.4,636,384, which is a continuation-in-part of U.S. patent applicationSer. No. 06/384,625, filed Jun. 3, 1982, now abandoned.

The contents of all of the above applications are all fully incorporatedherein by reference.

FIELD OF THE INVENTION

This invention is directed to the discovery of polypeptide fractions inmilk which contain peptides derived from allergens administered to amilk-producing animal. These fractions are useful in suppressingallergic responses in humans and animals.

BACKGROUND OF THE INVENTION

The allergic reaction in man and animals has been extensively studiedand the basic immune mechanisms involved are well known. The genericname for molecules that cause an allergic reaction is allergen. Thereare numerous species of allergens. Common examples include plantpollens, bee venom, house dust, animal dander, and a wide array of foodproteins. Many allergens are protein or polypeptide in nature, asproteins and polypeptides are generally more antigenic thancarbohydrates or fats. The allergic reaction occurs whentissue-sensitizing immunoglobulin of the IgE type reacts with foreignallergen. The IgE antibody is bound to mast cells and/or basophils, andthese specialized cells release chemical mediators of the allergicreaction when stimulated to do so by divalent antigens bridging theantibody molecules. Histamine, platelet activating factor, arachidonicacid metabolites, and serotonin are among the best known mediators ofallergic reactions in man.

The symptoms of the allergic reaction vary, depending on the locationwithin the body where the IgE reacts with the antigen. If the reactionoccurs along the respiratory epithelium the symptoms are sneezing,coughing and asthmatic reactions. If the interaction occurs in thedigestive tract, as in the case of food allergies, abdominal pain anddiarrhea are common. Systematic reactions, for example following a beesting, can be severe and often life threatening.

The preferred, but frequently impossible, method of relieving allergiesis allergen avoidance. Failing that, there are two medical approaches toallergy control, both with an approximate 60 to 85% efficacy rate (AasK., Allergy 37:1-14 (1982)). The most common approach to the medicaltreatment of allergies is to treat the symptom. Drugs known to block theeffects of the chemical mediators of the allergic reactions, includingantihistamines, are used to control the severity of the allergicsymptoms. These drugs, however, do nothing to prevent the allergicreaction and the liberation of the chemical mediators, and do nothing toprevent or diminish allergic responses to subsequent allergen exposure.

Another approach is to prevent the allergic reaction by desensitizingthe allergic host. This is accomplished by giving repeated small dosesof the reactive allergen. The treatment usually involves injecting theallergens under the skin. Treatment with reactive allergens, moreappropriately called immunotherapy, is believed to increase theconcentration of antibodies of the IgG type against the allergen. TheIgG antibody competes with the IgE antibody for allergen binding, andthis competitive antibody somehow neutralizes, arrests, or blocks theaction of the tissue sensitizing IgE antibody, although a distinctcorrelation between blocking antibody and amelioration of symptoms hasnot been definitely proven (Mailing, H. J., (ed.), ImmunotherapyPosition Paper, Allergy (Supp.) 6, 43:9-33 (1988)). This rationale,although generally accepted, is not fully understood, and does notreflect the complex interactions and events that accompany the IgGresponse. For example, other effects of immunotherapy that may beinvolved with relief of symptoms include suppression of IgE, increase inblocking IgA and IgG in secretions, reduced basophilreactivity/sensitivity, and reduced lymphocyte responsiveness toallergens. All of these changes may not occur in every patient, andthose related to actual symptom relief have not been conclusivelydefined (Norman, P. S., J. Allergy Clin. Immunol. 75:531-545 (1985)).

Immunotherapy using reactive allergen is dangerous because thesensitized host is actually treated with the molecules capable ofeliciting an allergic response. The treatment is started with extremelylow does to avoid inducing a severe reaction. The antigen concentrationrequired for 50% histamine release from peripheral basophils of anallergic individual varies 10,000 fold from patient to patient (Norman,P. S., J. Allergy Clin. Immunol. 75:531-545 (1985)). If no adversereaction occurs, higher doses are given. The injection may cause severeallergic reactions and extreme care must be taken. Only experienceddoctors can administer this treatment because if a severe reactionoccurs immediate medical treatment must be given to control the symptomsof the allergic reaction. Desensitization is an expensive, painful, andtime consuming process (Aas K., Allergy 37:1-14 (1982)), therefore, onlythe most severe allergies are treated by this method (Malling, H. J.,ed., Immunotherapy Position Paper, Allergy (Supp.) 6, 43:9-33 (1988)).

Modifying allergens to eliminate their allergenicity, or ability toinduce an IgE-mediated response, while preserving their immunogenicity,or their ability to elicit the protective IgG response, has been underinvestigation for years. Polymerized antigens have been evaluated withthe theory that they would display reduced antigenicity due to concealedantigenic determinants, a lower molecular concentration hence decreasedbridging opportunity on a weight basis, and slower diffusion through thetissues (Patterson R., J. Allergy Clin. Immunol. 68:85-90 (1981)).Allergen conjugates have been studied with a variety of allergens inattempts to selectively inhibit the formation of IgE antibodies whilenormal IgM and IgG responses to the antigen occurred (Lee, W. Y. et al.,Imm. Rev. 41:200-217 (1978)). Conjugates with a glutamic acid/lysinecopolymer may be acting specifically on IgE lymphocytes, on suppressorand/or helper T cells, or a combination of mechanisms may be in effect(Liu, F. T. et al., Proc. Natl. Acad. Sci. USA 76:1430-1434 (1979)).

Modifications that decrease the size of the allergen in efforts toeliminate their all ergenicity have also been attempted. It has beenproposed that limited proteolysis may actually reveal suppressordeterminants on allergen molecules (Mowatt A. M., Immunology 56:253-260(1985)).

Polypeptide fractions of allergens and the use of enzymes to break downthe parent proteins is not new (King, T. P., Advances in Immunology23:77 (1976)). U.S. Pat. No. 4,469,677 teaches the use of polypeptidefractions prepared from allergens for desensitization of allergic humansand animals. The polypeptide fractions are prepared by digestingallergens with proteolytic enzymes. Following the enzymatic digestionprocedure, the enzymes and residual parent allergens must be removedfrom the polypeptide subfractions. The specific structure of thepolypeptide fractions is dependent on the enzymes used for the digestiveprocess. However, this is a synthetic process that requirespre-selection of the enzymes used for the digestive process. Thepolypeptide fraction is produced by controlled proteolytic digestion ofthe polypeptide allergen. Although this represents an improvement overusing the reactive antigens, the treatment still requires frequent andpainful injections.

The function of the polypeptide fractions for suppressing immunefunction are structure dependent. see Unanue, E. R. et al., Science236:551-557 (1977) for a review of the structure dependence ofpolypeptide fractions of proteins in the immune response.

The structure of the polypeptide fractions of Michael is limited becausethe selection of enzymes is based on a limited understanding of whichenzymes are most important.

The development of oral "tolerance" is a normal phenomenon, and anecessary function in response to the variety of foreign antigensconsumed in the diet. Oral tolerance is initiated by a special class ofT lymphocytes and their products, and it results in systematicsuppression of the IgE-mediated hypersensitivity reaction. Variableresponses of other components of the immune system have been reported tooccur during induction of the tolerized state, so that the actualmechanism has not been succinctly defined.

In addition, animal studies have shown an actual enhancement of IgEproduction in response to intragastric administration of pollen extracts(Henderson, D. C. et al., Int. Archs. Allergy Appl. Immun. 79:66-71(1986)). Although this may be a dose-related phenomenon, conflictingresults were reported from two studies where 20 mg ovalbumin wasadministered orally to parenterally immunized mice, with IgE increasedin one study (Handson, D. G. et al., Int. Arch. Allergy Appl. Immun.55:256-532 (1977)) and decreased in the other (Lafont, S. et al., J.Exp. Med. 155:1573-1578 (1982)).

Some investigators have concluded that recognition of antigenicdeterminants varies between parenterally and orally induced suppressor Tcells. Suppressor cells resulting from feeding antigen were able torecognize different forms of the antigen, while parenterally inducedsuppressor T cells were specific for the molecular confirmation to whichthey initially responded (Mowatt, A. M., Immunology 56:253-260 (1985)).If this phenomenon is exhibited by a majority of antigens, it wouldfurther warrant the use of oral immunotherapy.

The problem with administering allergens orally to allergic subjects isthat severe immune reactions may occur following treatment by this routeas well. Anaphylaxis, relapse, urticaria, rectal bleeding, andanaphylactic shock have all been reported following oral immunotherapy(Platts-Mills, T. A. E., J. Allergy Clin. Immunol. 80:129-132 (1987)).In addition, oral treatment requires significantly larger doses thanparenteral therapy, due to the action of the digestive tract enzymesupon the allergens. Individual responses to the ingested antigens mayvary, however, so that more or less intact allergen may be representedto the gastrointestinal mucosa from the same dose given to differentsubjects, increasing the risk of inappropriate dosing.

Nonallergenic fractions of allergens can be given orally without concernfor severe immune reactions; however, for reasons that are not clearlyunderstood, oral administration of the polypeptide fractions of someallergens, including those prepared in the Michaels patent (U.S. Pat.No. 4,469,677) are not effective in causing desensitization. Studies ofthe immune response to feeding other modified proteins are limited, withvarying results. Collagen-induced arthritis in mice was suppressed byfeeding native type II collagen but not if the denatured molecule wasfed (Nagler-Anderson, C. L. et al., Proc. Natl. Acad. Sci. USA 83:7443(1986)). In a study of another autoimmune disease, both thedisease-producing and nondisease-producing fragments and decapeptides ofthe myelin base protein were capable of eliciting immune suppressionfollowing oral treatment in rats (Higgins, P. J. et al., J. Immunology140:440-445, (1988)). Results from a mouse study evaluating induction oforal tolerance with both natural and denatured ovalbumin identicalsuppression of delayed type hypersensitivity response to each form(Mowat, A. M., Immunology 56:253-260 (1985)).

One factor that may be involved in the failure of some molecularfragments to effectively induce immune tolerance is the low pH of thestomach, which may further modify the polypeptide fraction, abrogatingits ability to induce tolerance.

Another factor involved in the failure of orally dosed allergenfragments to induce desensitization to future exposure may be the actualnature of the fragment itself. In vitro synthesis of fragments requiresspecific enzymes and conditions that may or may not result in thepreparation of a biologically optimal formulation. The fragment may beeffective when it is acutely exposed to the immune system followingintravenous administration, but may be so susceptible to modificationthat any alterations occurring in the gastrointestinal tract before itsexposure to the appropriate immune system components may destroy itseffectiveness.

A need exists, therefore, for an oral formulation of allergen fractionsthat, when administered orally, is sufficiently active to inducetolerance but is not allergenic. A need also exists for methods ofproducing such products.

SUMMARY OF THE INVENTION

This invention embodies the discovery of a method of manufacture, methodof use, and product by process. The generic form of the product, byprocess of this invention, is milk that contains polypeptide fractionsof allergens. The specific product by process of this invention is themilk polypeptide fraction that contains polypeptide fragments ofspecific allergens.

Specifically, the invention is directed to a milk produced by animmunized milk-producing animal, such milk containing non-immunoglobulinpolypeptide fragments of the allergens used to immunize the animal.

The invention is further directed to a food product containing the milkof the invention or an active fraction of said milk.

The invention is further directed to an oral vaccine for administrationto an allergic subject for relief of the allergic response in suchsubjects, such vaccine containing the milk of the invention or an activefraction thereof.

The invention is further directed to a method for the production of animmune suppressive product comprising the immunization of amilk-producing animal and the collection of milk from such animal aftersuch animal reaches an immune state.

The invention is further directed to a method for the desensitization ofa subject to an allergen which method comprises administration of themilk of the invention, or an active fraction there, to such subject, inan amount and for a time sufficient to produce an immune suppressiveeffect.

The invention is further directed to pharmaceutical compositions whichprovide to a subject who is in need of desensitization to an allergen,the active, polypeptide fragment-containing fractions of the inventionat amounts which are efficacious in the treatment and suppression ofsuch subject's allergic responses and allergic reactions.

Description of the Preferred Embodiments

The invention provides an oral formulation for desensitization, an oralvaccine, which is of significant medical benefit to the subject whoingests it. By subject is meant a human or other animal in need ofallergy desensitizing according to the method of the invention. The oralvaccine of the invention can be self administered, is less painful, andless expensive then vaccines which must be injected. The subjectingesting the oral vaccine of the invention is exposed to the allergenin a derivatized, processed form which allows such subject to acquiretolerance to such allergen.

The milk of the invention, or an active fraction thereof, may beadministered in any form which retains the allergic response suppressiveproperties of the milk, either powdered or liquid. Any food productwhich incorporates the active fractions of such milk may also beprovided, for example, skim milk, yogurt and cheese.

By an "active fraction" of the milk of the invention is meant apreparation or composition which has been extracted from the milk of theinvention and which retains the beneficial, allergicreaction-suppression properties of the unextracted milk of the inventionwhich are due to the presence of derivatized, processed forms of theallergen used to immunize such animal providing such milk.

The derivatized, processed form of the allergen of the invention isfound in the milk of a milk-producing animal which has been immunizedagainst the allergen. The advantage of administering the polypeptidefraction of the invention to a subject rather than administering theunprocessed allergen itself is that, unlike the unprocessed allergen,the allergic response suppressive polypeptide fraction of the milk ofthe invention is nonallergenic. Desensitization of the subject whoingests the milk of the invention occurs but there is no risk of severeallergic reactions in such subject because the allergen has beenprocessed to a non-allergic form by the immune system in themilk-producing host.

The allergen which is used as a source of the immune suppressant activepolypeptides can be any allergen which is capable of provoking an immunesystem response in the mammalian animal which processes such allergen.If an allergen is not capable of inducing an immune system response inone mammalian species, another species may be used. In a preferredembodiment, the milk-producer bovine dairy cow is used to process theallergen. However, any mammalian species may be used, including humans,and animals in the ovid, porcine, and equine species, etc.

The allergen preparation may contain a mixture of allergens or only onespecific allergen. Examples of allergen mixtures which may be protectedagainst by the administration of the milk preparation of the inventioninclude mixtures of pollens of various trees and grasses, venoms fromanimals and insects, and food allergens. Examples of specific allergenswhich may be protected against by the administration of the preparationof the invention include, for example, ragweed, bee venom, and wheatprotein in the case of a food allergy.

The method of manufacture of the polypeptide fraction of this inventionproduces a unique subfraction of the parent allergen, such subfractionexhibiting immune suppressive activity when used to treat a hostallergic to such allergen. By "immune suppressive activity" is meant theability of a composition to lessen allergic symptoms or allergicresponses in an allergic subject who is administered such composition By"allergic symptoms" is meant physiological reactions or conditions whichare induced in an individual in response to that individual's beingexposed to substances which are allergic for that individual, such as,for example, allergy-induced congestion, headaches, breathing, itching,swelling, sneezing, wheezing, coughing, rhinorrhea, and decreasedability to smell and taste.

The milk composition of the invention, and polypeptide fractionsproduced therefrom according to the process of this invention, lack theallergenicity of the parent allergen (the allergen used to immunize themilk-producing animal) but possess the ability to desensitize a hostadministered such compositions.

It is not necessary that the milk-producing animal be induced to, ormaintained in, a hyperimmune state to produce the milk of the invention.Further, it is not necessary that the milk-producing animal be inducedto an antibody-producing state to produce the milk of the inventionbecause the active compositions of the invention do not rely onantibodies to provide their protective and suppressive activity. It isonly necessary that the allergen-administered animal be provided withtime sufficient to process such allergen into a form which is secretedin the milk of the animal.

Because immune cell processing of the allergen is one of the earlieststeps in the immune response, milk containing the active polypeptides ofthe invention may be obtained relatively soon after administering theallergen, for example, 24 hours to a week after allergen administration.Cows administered allergen according to the method described hereinprovide such active polypeptides in their milk 24 hours after allergenadministration.

Any amount of the allergen which results in the appearance of the activepolypeptides of the invention in the milk of such animal may beadministered to the milk-producing animal. If the amount of such activepolypeptides in the milk of the allergen-administered animal is too lowto provide efficacious benefits to a subject in need of desensitization,such active polypeptides may be provided to the subject in a moreconcentrated form. Such concentration is obtainable using techniquesknown in the art, for example, by salt precipitation, evaporation orlyophilization of the compositions of the invention.

Active polypeptide-containing preparations may be combined from morethan one allergen-administered animal, and from more than one species ofallergen-administered animal so as to provide the subject who is in needof desensitization with a composition containing a variety ofpolypeptide subfractions from different genetic heritages.

Although inducing a milk-producing animal to an antibody producing stateor to a hyperimmune state is not necessary, neither is it detrimental tothe milk of the invention and continuous production of milk containingthe active polypeptides of the invention may be achieved by repeatedlyadministering the allergen to the milk-producing host.

Without meaning to be held to this theory, since the processingmechanism of the immune system is not fully understood, it is thoughtthat the foreign protein or polypeptide is ingested or phagocytized byspecial cells of the immune system, the most noteworthy of which are themacrophage and neutrophil. After administration to a milk-producinganimal, enzymes within the immune cell break down or derivatize theprotein or polypeptide allergen into forms herein interchangeably termed"processed" forms or "subfractions" of the allergen.

The immune cell-produced subfractions are unique and cannot be producedoutside of the natural host. The uniqueness is based first on thestructure of the parent allergen and second on the combination ofenzymes contained in the mammalian host's allergen processing cells. Theenzyme combination which is expressed in the mammalian allergenprocessing cells are genetically determined; other species each have aunique combination of enzymes. For example peptide fractions ofallergens processed by the immune cells of the bovine species aresimilar, but not identical, to polypeptide fractions processed by theimmune cells of other species. It is because of this genetic specificitythat the polypeptides produced by the process of this invention areunique and different from polypeptides produced by any other method.

Special immune cells called helper cells assist the macrophage inpresenting the processed antigens to T and B lymphocytes. The passage ofthe allergen subfraction with and/or from the macrophage through theblood and across the mammary gland and into the milk may further modifythe structure of the allergen polypeptide found in milk. This naturalbiological method of processing environmental antigens and transportingnatural immune suppressants into the milk provides a mechanism wherebythe dairy cow transfers immune protective factors to the calf throughmilk. These milk immune factors protect the nursing young of suchanimals against allergic reactions. Nature, through the process ofevolution, has found a method for the mother of the species to samplethe environment for potential allergens. The immune system thentransforms the environmental allergens into the factors that protectagainst allergic reactions. The mother passes the immune protectivefactors to the nursing infant through milk. The essence of the discoveryof the invention is that man can utilize this natural biological processto transform known allergens into immune protective polypeptidefractions that have utility for preventing and treating allergicreactions.

The milk immune suppressive factor is a polypeptide subfraction found inmilk and not the antibodies found in milk. Although it is probable thatantibodies are useful in treating allergies, milk antibody is not theproduct of this invention.

The milk of animals treated by the process of this invention containsspecific and unique polypeptide subfractions of the allergens used totreat the host. The same milk lacks the intact or underivatized parentallergen. The uniqueness of these polypeptide fractions results fromspecific enzymatic breakdown by specialized immune cells of the host.Blood and mammary tissue enzymes may also play a role in modifying thechemistry of the allergens before entry into the milk. Milk containingthese unique nonreactive polypeptides is useful as an oral immunetreatment for inducing protection and/or tolerance when fed to otherspecies including man. The milk allergen polypeptide fractions, whenisolated from the milk, are useful for suppressing allergic reactionsagainst the representative parent allergens.

The allergen may be administered to the milk-producing animal by variousroutes of administration including oral, rectal, vaginal, intramuscularinjection, subcutaneous injection, intradermal injection, transcutaneousadministration, etc. While, any route of treatment can be used foradministering the allergen, the preferred route of treatment is byintramuscular injection of the allergen into the host species. Theallergen, upon entry into the body of the milk-producing animal, isprocessed by the immune system of the milk-producing host.

Treatment of the allergic individual or animal with the compositions ofthe invention effects protection in that individual or animal againstthe parent allergen without danger of side effects that may occur whenthe parent molecule is used. Examples of protein or polypeptideallergens that can be processed by the mammalian milk system of mammalsinclude pollen allergens of weeds, grasses and trees, animal venoms andpoisons, and various food allergens. In a preferred embodiment, thebovine milk system is used. Theoretically, any allergen can be processedby the mammalian, and especially by the bovine, macrophage system toproduce immune suppressive polypeptide subfractions.

The process of this invention is advantageous because it uses the fullspectrum of processing enzymes available in the allergen processing cellof the mammalian immune system. A second advantage of the process ofthis invention is that the full spectrum of polypeptide fractions iscontained in the immune suppressive milk product and the immunesuppressant allergen fractions, when contained in a natural milkvehicle, are orally active. The process of this invention, does notrequire selection of specific enzymes to degrade the allergen in vitroor regulation of the enzymatic digestive process, nor does the processof this invention require separation of the enzymes and residual parentmolecules from the reaction system. The entire antigen processing occursnaturally within the body of the milk-producing animal. The genetics ofthe mammal selects the enzymes and the natural metabolism of the mammalmaintains reaction conditions at an optimal level for antigenprocessing.

The peptide fractions of the invention may be monitored, purified andisolated by any technique or combination of techniques wherein suchpeptides retain their bioactivity, that is, wherein such peptides retainthe ability to suppress allergic responses. Such techniques are known inthe art and include, but are not limited to, chromatographic techniques,including adsorption chromatography, gel chromatography and especially,gel chromatography through a dextran, polyacrylamide, or agarose matrix,ion-exchange chromatography, for example, utilizing a strong cationicexchanger such as, for example, SP-Sephadex (which contains asulfopropyl functional group derivatized to a dextran), AG 50 (whichcontains a sulfonic acid derivatized to styrene-divinyl-benzene), andBio-Rex 40. (which contains a sulfonic acid functional group derivatizedto a phenolic matrix); weak cationic exchangers, such as, for example,CM-Sepharose (which contains a carboxymethyl functional groupderivatized to a dextran), Bio-Rex 70 (which contains a carboxylicfunctional group derivatized to a phenolic matrix); strong anionicexchangers, such as, for example, QAE-Sephadex (which contains adiethyl-(2-hydroxypropyl)-aminoethyl functional group derivatized to adextran), AG 1 (which contains a tetramethylammonium ion functionalgroup derivatized to styrene-divinyl-benzene); weak anionic exchangers,such as, for example, DEAE-Sephadex (which contains a diethylaminoethylfunctional group derivatized to a dextran, AG-3 (which contains atertiary amino functional group derivatized to an epoxyamine matrix);medium strength cationic exchangers, such as, for example, CM-cellulose(which contains a carboxymethyl functional group derivatized tocellulose), P-cel (which contains a phospho-functional group derivatizedto cellulose); and medium strength anionic exchangers, such as, forexample, DEAE-cellulose (which contains a diethylaminoethyl functionalgroup derivatized to cellulose), PEI-cellulose (which contains apolyethyleneimine functional group derivatized to cellulose,DEAE(BND)-cellulose (which contains a benzoylated-naphthoylated,diethylaminoethyl functional group derivatized to cellulose), PABcellulose (which contains a p-aminobenzyl functional group derivatizedto cellulose); and, exchangers which provide a mixture of functionalgroups, such as, for example, AG501 (which contains two functionalgroups, sulfonic acid and tetramethylammonium ion derivatized to astyrene-divinyl benzene matrix). In addition, high pressure liquidchromatography, may be used to separate the peptides of the invention.

Other compounds may be conjugated, either chemically or by geneticengineering, to the active polypeptides of the invention so as toenhance or provide additional properties to such polypeptide-containingcompositions, especially properties which enhance the activepolypeptide's ability to promote relief of an allergen's effects in asubject in need of immune suppressive treatment.

Amounts and regimens for the administration of the compositions of theinvention can be determined readily by those with ordinary skill.Generally, the dosage of the active peptide-containing composition willvary depending upon considerations such as: type of allergen employed;age of the subject being treated; health of the subject being treated;type of allergy being treated; kind of concurrent treatment, if any;physiological tolerance to the compositions of the invention; frequencyof treatment and the nature of the effect desired; gender; duration ofthe symptoms; and, counterindications, if any, and other variables asmay be adjusted, as desired, by the individual's physician. Dosage canbe administered in one or more applications to obtain the desiredresults.

The compositions of the invention can be administered in any appropriatepharmacological carrier for administration. They can be administered inany form that effects prophylactic, palliative, preventative or curingconditions of allergic reactions in humans and animals.

Preparations of the active polypeptide-containing compositions of theinvention for parenteral administration includes sterile aqueous ornon-aqueous solvents, suspensions and emulsions. Examples of non-aqueoussolvents are propylene glycol, polyethylene glycol, vegetable oil, fishoil, and injectable organic esters. Aqueous carriers include water,water-alcohol solutions, emulsions or suspensions, including saline andbuffered medical parenteral vehicles including sodium chloride solution,Ringer's dextrose solution, dextrose plus sodium chloride solution,Ringer's solution containing lactose, or fixed oils. Intravenousvehicles include fluid and nutrient replenishers, electrolytereplenishers, such as those based upon Ringer's dextrose and the like.

The compositions of the invention may also be administered by means ofpumps, or in sustained-release form.

Administration in a sustained-release form is more convenient for thepatient when continuous repeated administration for prolonged periods oftime are indicated.

The active polypeptide-containing compositions of the invention can beemployed in dosage forms such as tablets, capsules, powder packets, orliquid solutions for oral administration.

The pharmaceutical compositions of the present invention aremanufactured in a manner which is in itself know, for example, by meansof conventional mixing, granulating, dragee-making, dissolving,lyophilizing or similar processes.

Having now generally described the invention, the following examplesfurther describe the materials and methods used in carrying out theinvention. The examples are not intended to limit the invention in anymanner.

EXAMPLES Example 1

Immunization of Cows and Collection of Immune Milk

The immunogen for the preparation of the milk product for allergysymptom alleviation comprises a mixture of commercial allergen extractspurchased as 1:100 wt:vol preparations and pooled in the followingratios:

    ______________________________________    ALLERGEN         Volume (ml)    ______________________________________    Alternaria tenuis                     10    Aspergillus niger                     10    Monilia albicans 10    Hormodendrum hordei                     10    Lambs quarter    10    House dust       10    Helminthosporium Sat.                     10    Russian thistle  10    Careless weed    10    Spiny pigweed    10    Mugwort, common  10    Ragweed, slender 10    Ragweed, southern                     10    Ragweed, false   10    Ragweed, short   10    Ragweed, giant   10    Pigweed, redroot 10    Trichophyton mentag                     10    Goldenrod        10    Palmer's ameranth                     10    Sage, prairie    10    Curvularia spisifera                     30    Pullularia pullulans                     30    Mucor plumbeus   30    Fusarium moniliforme                     30    GS Mold Mix #2   30    GS 10 Tree Mix   30    Rhizopus nigricans                     30    ______________________________________

The pooled mixture is diluted with an equal volume of sterile saline.Cows are injected intramuscularly with 5 ml of the preparation at 2 weekintervals during normal lactation, and the milk is collected andpowdered. Milk is collected beginning 24 hr. post treatment.

Example 2

Fractionation of Milk Fractions Containing Processed Allergens

Twenty liters of fresh milk from hyperimmunized cows were run through acream separator (DeLaval Model 102) to remove the fat.

The resulting sixteen liters of skimmed milk was ultrafiltered to removethe high molecular weight species (over 100,000 daltons) using a hollowfiber diafiltration/concentrator. The concentrator is equipped with two100,000 daltons molecular weight cut-off cartridges. The skimmed milkwas run at the pump speed of 80 on the meter and inlet and outletpressure of 30 psi and 25 respectively.

Twelve liters of the filtrate (<100,000 daltons) coming out of thecartridges at the flow rate of four liters per hour was frozen orlyophilized for storage.

The same methods may be used to fractionate the milk fromnon-hyperimmunized cows.

Example 3

Desensitization of Allergic Subjects

Human volunteers consumed approximately 1/2 cup of the immune milkpowder reconstituted with 8 oz of water. Two hundred volunteers rangedin age from 4 to 87 years, and drank the milk for 1 to 172 months. Of168 subjects with allergies who submitted responses indicating thestatus of their condition, 138 (82.1%) indicated that they had improvedwhile drinking the milk. Ninety-four percent of subjects reportingbenefits stated their improvement occurred within 1 month of starting todrink the milk. Specific effects reported included less congestion,fewer or no headaches, easier breathing, less itching, swelling,sneezing, wheezing, coughing, rhinorrhea, and an enhanced ability tosmell and taste. Many subjects reported a recurrence of symptoms of milkconsumption was stopped, and several subjects discontinued allergyimmunotherapy after drinking the milk.

Having now fully described the invention, it will be understood by thosewith skill in the art that the scope may be performed within a wide andequivalent range of conditions, parameters and the like, withoutaffecting the spirit or scope of the invention or any embodimentthereof.

What is claimed is:
 1. A composition, wherein said composition comprisesan active non-antibody-containing fraction of milk, and wherein saidfraction is produced by the process comprising:(a) administering anallergen to a milk-producing animal; (b) collecting the milk from theanimal of part (a); (c) filtering the milk of part (b) through a filterwhich excludes molecules of greater than 100,000 daltons; and (d)collecting the effluent from the filtration of part (c) wherein saideffluent contains said active fraction.
 2. A method for the productionof an allergic response-suppressive pharmaceutical composition, whereinsaid method comprises:(a) administering an allergen to a milk-producinganimal; (b) collecting the milk from the animal of part (a); (c)filtering the milk of part (b) through a filter which excludes moleculesof greater than 100,000 daltons; and (d) collecting the effluent fromthe filtration of part (c) wherein said effluent contains said activefraction.
 3. The method of claim 2 wherein said milk-producing animal isa bovine.